For more than 150 years, since the synthesis of urea by Friedrich Wöhler in 1828, molecular chemistry has developed a vast array of highly sophisticated and powerful methods for the construction of more complex molecular structures. Beyond the molecular chemistry based on the covalent bond, there lies the field of supramolecular chemistry, aims to gain control over the intermolecular bond. Supramolecular species are characterized both by the spatial arrangement of their components and by the nature of the intermolecular bonds that hold these components together. They possess well-defined structural, conformational, thermodynamic and kinetic properties. Research has been focused on utilization of such confined spaces to manipulate reaction dynamics, properties of the encapsulated guest molecules. This research presented in this thesis is a consolidated account of photophysical and photochemical reactions carried in water-soluble macrocycles, cavitands and dynamic host systems such as dendrimers and micelles. With the aid of NMR (1D and 2D) spectroscopic techniques, the host-guest complex characterization is executed.

The thesis is divided into two major parts. The first part focuses on the localisation of probes in lipid/polymeric bilayers and in GM1 micelles. Included in this thesis is a new approach based on electronic energy transfer/migration (FRET/DDEM), which efficiently determines transversal positions of fluorescent molecules in lipid bilayers. This approach has been used to locate newly synthesized lipid probes in DOPC bilayers. The label was introduced at the end of sn-2 acyl chains of variable length. Analytical models accounting for FRET exist for a limited number of basic geometries. Here, a combination of FRET and Monte Carlo simulations enables the localisation of probes in bicelles and in bilayers containing pores, i.e. in lipid systems with variable curvature, or in non-homogenous lipid systems. This approach has been used to test whether conical-like fluorescence probes have an increased affinity to highly curved regions, which would enable preferential labelling of membrane pores. A simplified FRET model has been applied to localize 2-pyridones, a class of potential drugs, in GM1 micelles. Since the localisation of drugs within nanoparticles might influence the release kinetics and loading efficiency, knowledge about the drug location is highly relevant. It turned out that all derivatives were localised at the core-shell interface of GM1 micelles. The second part of the thesis focuses mainly on the estimation of lipid nanodomain size by means of FRET, which still remains the most powerful method in this field. Limitations of FRET in the determination of domain size have been explored. We showed that the limitations of FRET are mainly caused by a low probes affinity to either the liquid-ordered or liquid-disordered phase. In the continuing work we provided a detailed dynamic and structural study of crosslinker-triggered formation of nanodomains. Here, two different domains have been revealed, i.e. i) domains whose size grows with increasing amount of added cholera toxin (CTxB), and to which CTxB binds tightly; ii) domains formed in membranes containing a slightly increased amount of sphingomyelin (as compared to i) whose size does not change during titration by additional CTxB and to which CTxB binds less tightly. / Disertace je rozdělena do dvou hlavníchčástí. Prvníčást se zabývá lokalizací značek v lipidových/polymerních dvojvrstvách a v GM1micelách. V práci prezentujeme nový přístup založený na přenosu/migraci elektronické energie (FRET/DDEM), jež umožňuje efektivně určovat vertikální pozici fluorescenčních molekul uvnitř lipidové dvojvrstvy. Tato metoda byla použita k lokalizaci nově syntetizovaných lipidových značek značených na konci sn-2 acylového řetězce s různou délkou v DOPC dvojvrstvách. Analytické modely popisující FRET existují pouze pro limitovaný počet základních geometrií. Kombinace FRETu s Monte Carlo simulacemi nicméně umožňuje lokalizaci značek v bicelách a v dvojvrstvách obsahujících póry, tj. v lipidových systémech s proměnlivým zakřivením a v nehomogenních lipidových útvarech. Tento přístup umožnil např. zjistit, zda kuželovitětvarované značky mají zvýšenou afinitu k vysoce zakřiveným oblastem dvojvrstvy, což by umožnilo preferenční značení pórů. Lokalizovány byly rovněž tři deriváty 2-pyridonů(potencionálních léčiv) v GM1micelách za použití jednoduchého modelu zohledňujícího FRET mezi donory a akceptory nacházejícími se v micelách. Lokalizace léčiv v nanočásticích ovlivňuje kinetiku uvolňování (release kinetics) a množství látky solubilizované v micelách (loading efficiency). Druhá část se především zabývá určováním velikostí lipidových nanodomén pomocí FRETu, který stále zůstává nejvíce výkonnou metodou v této oblasti. Zkoumány byly limitace FRETu v určování lipidových nanodomén. Ukázalo se, že tato omezení jsou především způsobena nízkou afinitou značek buď k Lonebo k Ldfázi. V navazující studii jsme poskytnuli detailní dynamickou a strukturní studii formace nanodomén indukované crosslinkerem. Objevili jsme dva typy domén: a) domény, jejichž velikost se zvětšuje s rostoucím množstvím přidaného cholera toxinu (CTxB) a k nimž se CTxB váže pevně a b) domény vzniklé v membránách se zvýšeným množstvím sfingomyelinu (ve srovnání s a)), jejichž velikost se nemění během titrace dodatečným CTxB a k nimž se CTxB váže méně pevně. / This thesis has been elaborated within the framework of the Agreement on JointSupervision (co-tutelle) of an International Doctoral Degree Programmebetween Charles University in Prague, Czech Republic and the Department of Chemistry at Umeå University, Sweden.

Nanoparticle-based drug delivery systems (NDDS) have emerged as a promising strategy for formulation of anticancer drugs due to their ability to passively target solid tumors via exploitation of the enhanced permeation and retention effect. In particular, nano-sized block copolymer micelles (BCMs) have proven to be a viable delivery vehicle for hydrophobic anticancer drugs. To further enhance the specificity of BCMs towards cancer cells, extensive research has been focused on the formulation of actively targeted BCMs with tumor cell binding antigens conjugated to their surface. However, the in vivo transport of passively and actively targeted BCMs has only been studied to a limited extent. This thesis explores the potential and limitations of passively and actively targeted BCMs, as NDDS for delivery to solid tumors. The in vivo transport of BCMs at the whole body, tumor, and cellular levels is investigated in human breast cancer xenografts. Overall, active targeting of BCMs with epidermal growth factor (EGF) as the tumor cell binding antigen was not found to alter the whole body clearance of the vehicles; however, particle size had a profound effect on their pharmacokinetics and biodistribution profiles. Both passively and actively targeted BCMs exhibited heterogeneous distribution throughout solid tumors, with preferential localization in the tumor periphery and/or highly vascularized regions. In addition, the BCMs were found to exhibit impaired tumor penetration due to limited mobility and/or the binding site barrier. Although active targeting increases the in vivo BCM cellular uptake, the BCMs largely remained in the extracellular compartment, indicating that incomplete BCM delivery to all tumor cells remains as a major biological barrier. Interestingly, EGF-conjugated BCMs induced a potent bystander effect in vitro as a result of the paradoxical apoptotic effect of EGF, which has the potential to treat nearby tumor cells that do not respond directly to BCM treatment in vivo. In this way, EGF-BCMs may be beneficial for rendering the aforementioned in vivo barriers such as limited tumor penetration, as well as heterogeneity in tumor vascularization and receptor expression.

Nanoparticle-based drug delivery systems (NDDS) have emerged as a promising strategy for formulation of anticancer drugs due to their ability to passively target solid tumors via exploitation of the enhanced permeation and retention effect. In particular, nano-sized block copolymer micelles (BCMs) have proven to be a viable delivery vehicle for hydrophobic anticancer drugs. To further enhance the specificity of BCMs towards cancer cells, extensive research has been focused on the formulation of actively targeted BCMs with tumor cell binding antigens conjugated to their surface. However, the in vivo transport of passively and actively targeted BCMs has only been studied to a limited extent. This thesis explores the potential and limitations of passively and actively targeted BCMs, as NDDS for delivery to solid tumors. The in vivo transport of BCMs at the whole body, tumor, and cellular levels is investigated in human breast cancer xenografts. Overall, active targeting of BCMs with epidermal growth factor (EGF) as the tumor cell binding antigen was not found to alter the whole body clearance of the vehicles; however, particle size had a profound effect on their pharmacokinetics and biodistribution profiles. Both passively and actively targeted BCMs exhibited heterogeneous distribution throughout solid tumors, with preferential localization in the tumor periphery and/or highly vascularized regions. In addition, the BCMs were found to exhibit impaired tumor penetration due to limited mobility and/or the binding site barrier. Although active targeting increases the in vivo BCM cellular uptake, the BCMs largely remained in the extracellular compartment, indicating that incomplete BCM delivery to all tumor cells remains as a major biological barrier. Interestingly, EGF-conjugated BCMs induced a potent bystander effect in vitro as a result of the paradoxical apoptotic effect of EGF, which has the potential to treat nearby tumor cells that do not respond directly to BCM treatment in vivo. In this way, EGF-BCMs may be beneficial for rendering the aforementioned in vivo barriers such as limited tumor penetration, as well as heterogeneity in tumor vascularization and receptor expression.

Thesis (Ph. D.)--University of Missouri-Columbia, 1999. / Typescript. Vita. Includes bibliographical references. Also available on the Internet.

Thèse de doctorat : Sciences pharmaceutiques : Strasbourg 1 : 2008. / Titre provenant de l'écran-titre. Bibliogr. p. 321-327.

Reproduction de : Thèse de doctorat : Chimie et physico-chimie des polymères : Le Mans : 2008. / Titre provenant de l'écran-titre.

Thesis (M.Sc.)--Memorial University of Newfoundland, 2001. / Restricted until June 2002. The CD-ROM "contains the latest version of the simulation code. The code is contained in .tar.gz, .tar.Z and .zip files. There are also brief instructions for the interested reader on how to compile and run the simulations"--Appendix A. Bibliography: leaves 122-137.

Thesis (Ph. D.)--University of Missouri-Columbia, 1999. / Typescript. Vita. Includes bibliographical references. Also available on the Internet.

Les micelles polyioniques ont émergé comme des systèmes prometteurs de relargage de médicaments hydrophiles ioniques. Le but de cette étude était le développement des micelles polyioniques à base de dextrane pour la relargage de médicaments hydrophiles cationiques utilisant une nouvelle famille de copolymères bloc carboxymethyldextran-poly(éthylène glycol) (CMD-PEG). Quatre copolymères CMD-PEG ont été préparés dont deux copolymères identiques en termes de longueurs des blocs de CMD et de PEG mais différent en termes de densité de charges du bloc CMD; et deux autres copolymères dans lesquels les blocs chargés sont les mêmes mais dont les blocs de PEG sont différents. Les propriétés d’encapsulation des micelles CMD-PEG ont été évaluées avec différentes molécules cationiques: le diminazène (DIM), un médicament cationique modèle, le chlorhydrate de minocycline (MH), un analogue semi-synthétique de la tétracycline avec des propriétés neuro-protectives prometteuses et différents antibiotiques aminoglycosidiques. La cytotoxicité des copolymères CMD-PEG a été évaluée sur différentes lignées cellulaires en utilisant le test MTT et le test du Bleu Alamar. La formation de micelles des copolymères de CMD-PEG a été caractérisée par différentes techniques telles que la spectroscopie RMN 1H, la diffusion de la lumière dynamique (DLS) et la titration calorimétrique isotherme (ITC). Le taux de relargage des médicaments et l’activité pharmacologique des micelles contenant des médicaments ont aussi été évalués. Les copolymères CMD-PEG n'ont induit aucune cytotoxicité dans les hépatocytes humains et dans les cellules microgliales murines (N9) après 24 h incubation pour des concentrations allant jusqu’à 15 mg/mL. Les interactions électrostatiques entre les copolymères de CMD-PEG et les différentes drogues cationiques ont amorcé la formation de micelles polyioniques avec un coeur composé du complexe CMD-médicaments cationiques et une couronne composée de PEG. Les propriétés des micelles DIM/CMDPEG ont été fortement dépendantes du degré de carboxyméthylation du bloc CMD. Les micelles de CMD-PEG de degré de carboxyméthylation du bloc CMD ≥ 60 %, ont incorporé jusqu'à 64 % en poids de DIM et ont résisté à la désintégration induite par les sels et ceci jusqu'à 400 mM NaCl. Par contre, les micelles de CMD-PEG de degré de carboxyméthylation ~ 30% avaient une plus faible teneur en médicament (~ 40 % en poids de DIM) et se désagrégeaient à des concentrations en sel inférieures (∼ 100 mM NaCl). Le copolymère de CMD-PEG qui a montré les propriétés micellaires les plus satisfaisantes a été sélectionné comme système de livraison potentiel de chlorhydrate de minocycline (MH) et d’antibiotiques aminoglycosidiques. Les micelles CMD-PEG encapsulantes de MH ou d’aminoglycosides ont une petite taille (< 200 nm de diamètre), une forte capacité de chargement (≥ 50% en poids de médicaments) et une plus longue période de relargage de médicament. Ces micelles furent stables en solution aqueuse pendant un mois; après lyophilisation et en présence d'albumine sérique bovine. De plus, les micelles ont protégé MH contre sa dégradation en solutions aqueuses. Les micelles encapsulant les drogues ont maintenu les activités pharmacologiques de ces dernières. En outre, les micelles MH réduisent l’inflammation induite par les lipopolysaccharides dans les cellules microgliales murines (N9). Les micelles aminoglycosides ont été quant à elles capable de tuer une culture bactérienne test. Toutefois les micelles aminoglycosides/CMDPEG furent instables dans les conditions physiologiques. Les propriétés des micelles ont été considérablement améliorées par des modifications hydrophobiques de CMD-PEG. Ainsi, les micelles aminoglycosides/dodecyl-CMD-PEG ont montré une taille plus petite et une meilleure stabilité aux conditions physiologiques. Les résultats obtenus dans le cadre de cette étude montrent que CMD-PEG copolymères sont des systèmes prometteurs de relargage de médicaments cationiques. / Polyion complex (PIC) micelles have emerged as promising delivery systems of ionic hydrophilic drugs. It was the aim of this study to develop dextran-based PIC micelles for the delivery of hydrophilic cationic drugs using a new family of carboxymethyldextranblock- poly(ethylene glycol) (CMD-PEG) copolymers. Four CMD-PEG copolymers were prepared: (i) two copolymers identical in terms of the length of CMD and PEG blocks, but different in terms of the charge density of the CMD block; and (ii) two copolymers in which the charged block is the same, but the PEG block is of different molecular weight. The micellization of CMD-PEG copolymers and drug delivery aspects of the resulting micelles were evaluated using different cationic drugs: diminazene (DIM), a model cationic drug, minocycline hydrochloride (MH), a semisynthetic tetracycline antibiotic with promising neuroprotective properties and different aminoglycoside antibiotics. The cytotoxicity of CMD-PEG copolymers was evaluated in different cell lines using MTT and Alamar blue assays. CMD-PEG micelles encapsulating different drugs were characterized using different techniques, such as 1H NMR spectroscopy, dynamic light scattering (DLS), and isothermal titration calorimetry (ITC). The pattern of drug release and pharmacological activity of micelles-encapsulated drugs were also evaluated. The CMD-PEG copolymers did not induce cytotoxicity in human hepatocytes and murine microglia (N9) in concentrations as high as 15 mg/mL after incubation for 24 h. Electrostatic interactions between CMD-PEG copolymers and different cationic drugs triggered the formation of PIC micelles with a CMD/drug core and a PEG corona. The properties of DIM/CMD-PEG micelles were strongly dependent on the degree of carboxymethylation of the CMD block. Micelles of CMD-PEG copolymers having degree of carboxymethylation ≥ 60%, incorporated up to 64 wt% DIM, resisted salt-induced disintegration in solutions up to 400 mM NaCl and sustained DIM release under physiological conditions (pH 7.4, 150 mM NaCl). In contrast, micelles of CMD-PEG of degree of carboxymethylation ~ 30% had lower drug content (~ 40 wt% DIM) and disintegrated at lower salt concentration (∼ 100 mM NaCl). The CMD-PEG copolymer that showed the most satisfactory micellar properties, in terms of high drug loading capacity, sustained drug release and micelles stability was selected as a potential delivery system of minocycline hydrochloride (MH) and different aminoglycosides. CMD-PEG micelles encapsulating either MH or aminoglycosides had small size (< 200 nm in diameter), high drug loading capacity (≥ 50 wt% drug) and sustained drug release. These micelles were stable in aqueous solution for up to one month, after freeze drying and in the presence of bovine serum albumin. Furthermore, the micelles protected MH against degradation in aqueous solutions. Micelles-encapsulated drugs maintained their pharmacological activity where MH micelles reduced lipopolysaccharides-induced inflammation in murine microglia (N9) cells. And aminoglycosides micelles were able to kill a test micro-organism (E. coli X-1 blue strain) in culture. Aminoglycosides/CMD-PEG micelles were unstable under physiological conditions. Micelle properties were greatly enhanced by hydrophobic modification of CMD-PEG. Thus, aminoglycosides/dodecyl-CMD-PEG micelles showed smaller size and better stability under physiological conditions. The results obtained in this study show that CMD-PEG copolymers are promising delivery systems for cationic hydrophilic drugs.